Proper functioning of the immune system, in detecting and mounting an immune response against invading pathogens, is dependent on lymphocyte recirculation through secondary lymphoid organs. Factors that affect lymphocyte entry and exit into these lymphoid tissues are not completely understood. Furthermore, persistent pathological disruption of lymphocyte recirculation can lead to sequestration of lymphocytes in lymphoid organs, as seen in HIV infection, and may contribute to immunosuppression of the host. The long-term goal of our research program is to gain a detailed molecular understanding of regulation of lymphocyte trafficking in acute and chronic inflammation using viral infections as a model system. We have found that expression of a phospholipid receptor, sphingosine-1-phosphate receptor-1 (S1P1), within lymphocytes is required for their ability to exit from secondary lymphoid organs. Both innate and adaptive immune responses alter the expression and function of this receptor leading to lymphocyte sequestration. A variety of cytokines, including interferon alpha/beta (IFN), up-regulate expression of the transmembrane C-type lectin, CD69. CD69 interacts with and down-modulates surface expression and function of S1P1, a G protein coupled receptor, leading to transient lymphocyte sequestration. In addition, engagement of the antigen receptor on T cells leads to transcriptional down-regulation of S1P1 mRNA expression and a prolonged inhibition of lymphocyte egress. Thus, S1P1 function is tightly regulated by two distinct mechanisms during the early stages of an immune response. The goal of this proposal is to further understand the regulation of lymphocyte trafficking by S1P1 and CD69 and its impact on generation of T cell responses. Cellular and biochemical approaches will be utilized to determine: (1) the molecular requirements for S1P1-CD69 interaction;(2) the molecular mechanism of inhibition of S1P1 function by CD69;and (3) the effects of genetic perturbation of regulation of S1P1 expression and function on in vivo T cell responses to a viral infection. Our hope is that a detailed understanding of factors that regulate lymphocyte trafficking can contribute not only to vaccine development and improved immunotherapies, but also provide targets for therapeutic disruption of these processes in autoimmune and chronic inflammatory diseases. PUBLIC HEALTH RELEVENCE: Circulation of white blood cells through lymphoid tissues is essential for initiation of immune responses to infectious microorganisms such as viruses but could also have detrimental consequences in chronic inflammatory and autoimmune diseases. The goal of this proposal is to gain a detailed understanding of how two specific molecules known to affect white blood cell trafficking regulate this process. The knowledge gained from these studies will facilitate development of therapies that target white blood cell trafficking for treatment of inflammatory diseases.